The two pore domain potassium channels (K2p's) are divided into six subfamilies: TWIK, TREK, TASK, TALK, THIK, and TRESK. They are formed by dimerization of subunits that have two tandem pore domains each resembling that of the voltage-gated potassium channels. TASK3 is a member of the TASK subfamily, and has proposed functions of sensing metabolic level, conferring anesthetic sensitivity, as well as modulating neuronal excitability and cancer cell proliferation, based on early studies on tissue distribution, electrophysiology, and manipulative intervention with non-specific reagents. Therefore, TASK3 has been considered a potential therapeutic target for cancer and neurological disorders. However, due to the lack of selective ligands, progress in the study on this channel has been very slow. We propose to perform a high-throughput screen for compounds that specifically target TASK3 channels, and further develop them into small molecular probes. In our preliminary studies, we have constructed a HEK-293 cell line expressing TASK3, and optimized a Tl+ - based fluorescence assay in 384-well format on FDSS. In addition, we have also developed secondary screens with electrophysiology and counter screens using wild-type HEK-293 cell line, and HEK-293 expressing THIK-2 and other potassium channels. A validation screen using the LOPAC library from Sigma- Aldridge has led to the identification of a compound, N1786, that inhibits TASK3. Small molecular probes that specifically interact with TASK3 may become valuable tools for studying the structure and functions of these unique ion channels, and developing targeted therapeutic approaches. PUBLIC HEALTH RELEVANCE: The two-pore-domain potassium channels (K2p's) play important roles in modulating excitability, sensing environmental conditions, as well as regulating cellular secretion and proliferation. In particular, TASK3 has been considered a potential therapeutic target for cancer and neurological disorders. However, very little is known about these channels, because of the lack of specific ligands. We propose to perform a high-throughput screen for compounds that specifically target TASK3 channels, and further develop them into small molecular probes.